STRUCTURE OF THE O-GLYCOPEPTIDES ISOLATED FROM BOVINE-MILK COMPONENT PP3

The heat-stable acid-soluble phosphoglycoprotein component PP3 was iso lated from the bovine milk proteose peptone fraction by concanavalin A affinity chromatography. Glycopeptides from the ConA-bound fraction c orresponding to the component PP3 were obtained by Pronase digestion a nd were separated by gel filtration into high and low-molecular-mass g lycopeptides. In a previous work, we have investigated the structure o f the N-glycans from the high-molecular-mass glycopeptides [Girardet e t al. (1995) Eur J Biochem 234: 939-46]. Here, we describe the structu re of the O-glycans from the low-molecular-mass glycopeptides. By comb ining methylation analysis, mass spectrometry, 400 MHz H-1-NMR spectro scopy and peptide sequence analysis, we show that the low-molecular-ma ss fraction contains several neutral glycopeptides. A mixture of the f ollowing three glycan structures linked to the Thr(86) has been identi fied: GalNac alpha 1-O-Thr, Gal(beta 1-3)GalNAc alpha 1-O-Thr and Gal( beta 1-4)GlcNAc(beta 1-6)[Gal(beta 1-3)]GalNAc alpha 1-O-Thr.